An Alternative Metabolic/Antidotal Pathway Of All-trans-retinal In The Retina

Purpose:To investigate the role for formation of all-trans-retinal dimer(at1RAL-dimer)in the retina.Methods:The formation of atRAL-dimer in primary porcine retinal pigment epithelium(RPE)cells induced by all-trans-retinal(atRAL)was examined by high-perfonmance liquid chromatography(HPLC).Cellular toxicities of atRAL-dimer after 5 min irradiation(220 and 10000 lux)by LED light were tested by MTS assay at 6 h,24 h,3 days and 5 days.Moreover,crystal violet staining assav was adopted to compare the cytotoxicities of atRAL and atRAL-dimer.Photosensitivity of atRAL-dimer with atRAL and A2E under 220 and 10000 lux LED light was compared.Reaction mixtures of atRAL with porcine RPE/choroid,neural retina or photoreceptor outer segment(POS)were analyzed by HPLC.atRAL-dimer was irradiated by LED light(220 and 10000 lux)for 5 min,and analyzed by atmospheric pressure chemical ionization-mass spectrometry(APCI-MS).Results:After treatment of primary porcine RPE cells with atRAL,atRAL-dimer readily formed,and accumulated in a concentration-and time-dependent manner yet A2E was barely detected.Cell-based assays revealed that atRAL.the precursor of atRAL-dimer.significantly altered the nmorphology of primary porcine RPE cells and decreased cell viability at a concentration of 80 μM regardless of light exposure.By contrast atRAL-dimer was not cytotoxic and phototoxic to primary porcine RPE cells.Compared to atRAL and A2E,atRAL-dimer was more vulnerable to light.followed by the generation of its photocleaved products.Moreover,we observed the presence of-atRAL-dimer in reaction mixtures of atRAL with porcine POS-RPE/choroid or neural retina’Conclusions:We proposed an alternative metabolic/antidotal pathway of atRAL in RPE cells:atRAL that evades the participation of the visual(retinoid)cycle undergoes a condensation reaction to yield atRA L-dieter in both POS and RPE.Translocation of atRaL.all-trans N-retinylidene-phosphatidylethanolamine(NR-PE),atRAL-dimer and photocleavage products of atRAL-dimer from POS into the RPE is accomplished by phagocytizing shed POS on a daily basis.Without causing damage to RPE cells,light breaks up total atRAL-dimer within RPE cells to release low-molecular-weight photocleavage fragments.The latter,together with POS-atRAL-dimer photocleavage products,may easily move across membranes and thereby be metabolically eliminated.